[Preparation and characterization of a single chain antibody fragment of mAb SZ-21 against platelets GPIIIa].
摘要
OBJECTIVE: To prepare a single chain antibody (ScFv) of mAb SZ-21 against platelet GPIIIa for its future clinical application. METHODS: The expression vector pET20b-SZ-21ScFv was constructed and the fusion protein was expressed in E. coli BL21 (DE3) PlysS. The activated fusion protein was obtained after a series of purification steps, including cell breakage, inclusion body solubilization, His-bind resin affinity chromatography and protein refolding. RESULTS: The fusion protein yields were up to 21% of the total amount of bacteria protein. The ScFv fragment could inhibit ADP-induced platelets aggregation in a dose-dependent manner in vitro and the maximal inhibition rate was obtained at a concentration of 20 micro g/ml. It also reacted with endothelial cells as detected by flow cytometry. Moreover, the ScFv fragment was able to inhibit the binding of fibrinogen to platelet. CONCLUSION: The SZ-21ScFv fragment had the activity to inhibit platelets aggregation and the binding of fibrinogen to platelet, being potentially useful for the treatment of thrombotic diseases.